Project Summary Abstract: Repeated exposure to a drug of abuse induces alterations in neuronal activity and neuroplastic changes in multiple brain areas and circuits related to reward, impulse control and affect. These widespread changes contribute to multiple behavioral alterations. Despite differing in their molecular targets, different drugs cause common alterations in the brain. Increasing our knowledge of the impact of drugs of abuse on multiple brain regions and circuits is crucial to better understand the mechanisms that mediate the transition from drug use to addiction and facilitate the development of novel treatment. We hypothesize that phencyclidine induces neurotransmitter switching in more than just one brain region and that different categories of drugs induce a shared transmitter switch that is relevant for the transition from drug intake to addiction. To test these hypotheses we are developing genetic mouse models to facilitate high- throughput, whole-brain screening of transmitter switching involving GABA and glutamate. The current assays of transmitter switching involve time consuming sectioning of a single parts of the brain, staining sections and stereology, putting whole-brain analysis out of reach. The combination of genetically encoded fluorescent reporter proteins, brain clarification, light sheet microscopy and whole-brain automatic image analysis will speed analysis and accelerate understanding of the role of transmitter switching in drug-induced neuroplasticity and drug-induced behavioral alteration. These tools will be used to determine whether two different substances of abuse that have distinct targets in the brain, PCP and methamphetamine (METH), induce NTS in multiple brain regions and whether some PCP- and METH-induced NTS are the same.